Crystalline forms(2s)-n-5[amino(imino)methyl]-2-thienylmethyl-1-(2r)-2[(carboxymethyl) amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide nh2o

ABSTRACT

The present invention relates to crystalline forms of (2S)-N-5-[amino(imino)methyl]-2-thienyl-methyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide nH 2 O.

TECHNICAL FIELD

The present invention relates to crystalline forms of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide.nH₂Orepresented by the following Formula (1):

wherein n is the number of combined water per molecule and represents 0,1, 3, 4, 6 or 7.5.

BACKGROUND OF THE INVENTION

The free compound of Formula (1), i.e., compound to which acids were notadded, and pharmaceutically acceptable salts, hydrates, solvates, andisomers thereof are the subjects of Korean Patent Laid-Open PublicationNo. 2000-047461 and WO 0039124, and may be effectively used as newthrombin inhibitors.

The physical property of a drug has a huge effect on production anddevelopment process of its raw drug and development process of its finalproduct. A drug may be roughly divided into crystalline form andamorphous form according to its crystallinity. Some drugs may beobtained in both crystalline form and amorphous form, while other drugsmay be obtained only in either crystalline form or amorphous form.Crystalline form and amorphous form may exhibit large difference inphysicochemical properties. For instance, there is a report that an oralabsorption rate or bioavailability is different in some drugs becausesolubility and dissolution rate are different depending on whether thedrugs are in crystalline form or amorphous form (see, PharmaceuticalSolids: A Strategic Approach to Regulatory Considerations, Stephen Byrnet al, Pharmaceutical Research, 945, 12(7), 1995). Bioavailability of adrug is directly related to its effect and side effect. In other to say,to obtain the desired effect of a drug, a certain desired bloodconcentration should be reached. If the blood concentration becomesunduly high, a side effect or toxicity is accompanied. Bioavailabilitymay be improved by selecting a suitable crystalline form. Thus, thecrystalline form of a drug should be identified in the course ofdevelopment and approval of the drug.

Except special cases, it is easy to obtain a drug having crystallinityin the process of its research and development. A report shows that thecrystallinity of a drug may be an important advantage because in thefinal step for producing the drug, the drug may be purely obtainedthrough recrystallization that is a relatively easy purificationprocess, and a drug having crystallinity, whose physicochemicalproperties may be easily identified, is advantageous even in the qualitycontrol of its product process (see, An integrated approach to theselection of optimal salt form for a new drug candidate, Abu T. M.Serajuddin et al, International Journal of Pharmaceutics, 209, 105,1994). On the other hand, some drugs having crystallinity may havepolymorphism. An article reported that generally speaking, in case thatthe crystalline structure of a drug is different, its solubility orother physical properties may be different, and the crystalline form ofa drug may be changed under certain conditions [Pharmaceutical Solids: AStrategic Approach to Regulatory Considerations, Stephen Byrn et al.,Pharmaceutical Research, 945, 12(7), 1995]. Therefore, in case that adrug has polymorphism, to obtain purely all crystalline forms of thedrug and to discover physical properties of each form are very importantin the development and production of the drug.

BRIEF SUMMARY OF THE INVENTION

Accordingly, the present inventors have found crystalline forms usefulas thrombin inhibitors by obtaining various crystalline forms from thefree compound of the above Formula (1) and identifying their physicalproperties.

Therefore, the purpose of the present invention is to providecrystalline forms of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide.nH₂Orepresented by the following Formula (1):

wherein n is the number of combined water per molecule and represents 0,1, 3, 4, 6 or 7.5.

BRIEF DESCRIPTION OF DRAWINGS

FIG. 1 is a powder X-ray diffraction diagram of the crystalline Form Iof(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide.

FIG. 2 is a powder X-ray diffraction diagram of the crystalline Form IIof(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide.

FIG. 3 is a powder X-ray diffraction diagram of the crystalline Form IIIof(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide.

FIG. 4 is a powder X-ray diffraction diagram of the crystalline Form IVof(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide.

FIG. 5 is a powder X-ray diffraction diagram of the crystalline Form Vof(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide.

FIG. 6 is a powder X-ray diffraction diagram of the crystalline Form VIof(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide.

DETAILED DESCRIPTION

The free compound of the above Formula (1) may be prepared according toa known method (see, Korean Patent Laid-Open Publication No. 2000-047461and WO0039124).

The crystalline forms of Formula (1) of the present invention obtainedfrom the above free compound or other crystalline forms exist in theform of anhydride or hydrates having various combined water. Preferably,according to the recrystallization method and the number of combinedwater, the crystalline Form I (n=7.5), the crystalline Form II (n=4),the crystalline Form III (n=6), the crystalline Form IV (n=3), thecrystalline Form V (n=0), and the crystalline Form VI (n=1) may beobtained. For instance, the crystalline Form IV may be obtained bydissolving the free compound of Formula (1) in the mixed solvent ofwater, and methanol or ethanol while heating and recrystallizing it.

The crystalline Form V may be obtained by drying the crystalline Form IVunder vacuum. The crystalline Form VI may be obtained by moistureabsorption of the Form V. However, the Form I may be obtained bystirring the Form VI in water. The crystalline Form II may be obtainedby drying the Form I under vacuum. And, the Form III may be obtained bymoisture absorption of the Form II. Since the molecular weight of theabove free compound is 533.65, the theoretical water contents of thesehydrates of Formula (1) are 0, 3.3, 9.2, 11.9, 16.8, and 20.2%, to thehydrates of Formula (1) wherein n is 0, 1, 3, 4, 6, and 7.5,respectively. However, it is usual that the water contents of actuallyobtained samples deviate from the above theoretical values depending ondrying condition and drying time in preparation, amount of the surfacemoisture absorbed at the surface, etc. Therefore, the water content ofthe hydrate of Formula (1) wherein n is 0, i.e., anhydride of Formula(1), may be 0˜3%, that of the hydrate wherein n is 1 may be 2˜9%, thatof the hydrate wherein n is 3 may be 4˜11%, that of the hydrate whereinn is 4 may be 9˜15%, that of the hydrate wherein n is 6 may be 12˜20%,and that of the hydrate wherein n is 7.5 may be 16˜26%. Thus, toidentify the crystalline form of Formula (1), the water content shouldbe identified, with conducting the powder X-ray diffraction test.

Each crystalline form may be distinguished by characteristic peaks shownat the powder X-ray diffraction test. For example, as shown in Tables 1,2, 4, 5, 6, and 7, the crystalline Form I has characteristic peaksdistinguished from other crystalline forms at 7.3°, 9.1°, 18.0°, and28.8°, the crystalline Form II at 7.0°, 12.2°, 19.2°, and 20.0°, thecrystalline Form III at 10.6°, 19.4°, 20.9°, 21.6°, and 24.4°, thecrystalline Form IV at 10.0°, 16.7°, 20.8°, 21.9°, and 26.0°, thecrystalline Form V at 15.8°, 18.3°, 20.3°, 20.8°, and 26.5°, thecrystalline Form VI at 13.6°, 14.7°, 23.2°, and 27.5°. Further, as shownin FIGS. 1 to 6, it can be confirmed in the power X-ray diffractiondiagram that each crystalline form above has a different crystalstructure from one another.

A crystalline form may be changed according to storage condition such asrelative humidity, etc. Thus, it is important to confirm stability of acrystalline form according to storage condition. Among the abovecrystalline forms, the crystalline Form VI was identified as a stablehydrate whose crystal structure is not changed under any relativehumidity.

Karl-Fischer titrimetry has been widely used for determining the watercontent in samples (see, Quantitative Chemical Analysis, 4th edition, I.M. Koltmoff et al, 858, The Macmillan Company, 1969). When Karl-Fischertitrimetry was applied to the above crystalline forms, the water contentof the crystalline Form VI was proven as 3.5%, which corresponds to theweight ratio of a water molecule when n of Formula (1) is 1. On theother hand, the water content of the crystalline Form 1 was proven as20.2%, which corresponds to the weight ratio of a water molecule when nof Formula (1) is 7.5.

Moisture included in a sample is not completely removed even if thesample is dried under vacuum. In order to remove moisture completely,various drying agents should be placed with the sample under vacuum.Various kinds of drying agents may be used for the present invention:calcium sulfate, sodium sulfate, calcium chloride, etc. The most widelyused drying agent is P₂O₅ (see, MIT Laboratory techniques manual, MITdept. of Chemistry, 10:43, 1979). If the crystalline Form I is driedunder vacuum in a desiccator in which P₂O₅ is used as a drying agent,the moisture included in the crystalline form can be removed. Then, itis confirmed by the power X-ray diffraction test that the crystallineform was changed, and the changed form is identified as the crystallineForm II. The crystalline Form II became stable by adsorbing moisture andits water content is 10.8% that corresponds to the weight ratio of 4water molecules. If the crystalline Form II is left under highlyrelative humidity, the form is changed to the crystalline Form III, andits water content is 16.9% that corresponds to the weight ratio of 6water molecules.

From the above results, it can be seen that the crystalline Form I, FormII and Form III are hydrates wherein n is 7.5, 4, and 6, respectively.

The solvent to be used in recrystallization may be usually availablekinds of alcohols, which are alkanes alcohols having the carbon numberof 1 to 8, such as methanol, ethanol, propanol, butanol, isopropanol andoctanol, etc., but methanol and ethanol are preferable, and methanol isthe most preferable, but not limited to them. Furthermore, as a solventto be used to recrystallize the above free compound, in addition toalcohols exemplified above, water and organic solvents, such asn-hexane, ethylacetate, butylacetate, acetonitril, chloroform,diethylether, acetone, etc., and other usually available solvents may beused. The above free compound may be dissolved or dissolved in heating,by using one solvent or more than one in mixture among the above and maybe recrystallized.

If the above several crystalline forms are dissolved in alcohols, asuitable amount of water is added thereto, and the mixture isrecrystallized, the crystalline Form IV, another crystalline form, maybe obtained. The X-ray crystal structure method identified thecrystalline form as hydrate wherein n is 3. The crystalline Form IV wasdried under vacuum in the presence of P₂O₅ to obtain the crystallineForm V which is anhydride. The crystalline Form V is changed into thecrystalline Form VI by absorbing moisture. The crystalline Form VI has3.5% of water content, and is stable hydrate wherein n is 1.

The stress stability test showed that the crystalline form of thecompound of Formula (1) above is physicochemically more stable than theamorphous form. The amorphous form showed a residual content of only 87%as well as discoloration after 4 weeks' storage, especially at 70° C.However, the crystalline Form I and IV were stable withoutdiscoloration.

As Korean patent Laid-Open Publication No. 2000-047461 and WO0039124 aredisclosed, the free compound of Formula (1) of the present invention iseffectively used as a thrombin inhibitor. And, its crystalline forms arealso useful as thrombin inhibitors.

Below, the present invention will be explained in more detail withreference to the following examples, comparative examples, and testexamples. However, it should be understood that these examples have beendescribed as preferred specific embodiments of the present invention,and are not intended to limit the scope of the present invention in anyway. Other aspects of this invention will be apparent to those skilledin the art to which the present invention pertains.

EXAMPLES Example 1 Preparation of the crystalline Form II of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide

The crystalline Form I prepared in the following example 8 was driedunder vacuum in the presence of P₂O₅ for one day and then placed at therelative humility of 75% for one day to obtain the titled crystallineForm II.

Example 2 Preparation of the crystalline Form III of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide

The crystalline Form II prepared in Example 1 above was placed at therelative humidity of 93% for one day, and then moved and placed at therelative humidity of 64% for one day to obtain the titled crystallineForm III.

Example 3 Preparation of the crystalline Form IV (1) of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide

The free compound (1 g) of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-((carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamidewas placed into a glass container and then methanol (5.0 ml) was addedthereto. While stirring, the mixture was heated to obtain a clearsolution. Water (0.5 ml) was added to the solution and then the solutionwas cooled at room temperature. White crystals were obtained therefrom.The crystals were filtered and then washed with water. They were driedin the air (0.85 g, yield 85%).

Example 4 Preparation of the crystalline Form IV (2) of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamideThe free compound (1 g) of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamidewas placed into a glass container and dissolved by adding methanol (6.0milliliter), water (1.5 milliliter), and 6N hydrochloric acid solution(0.65 milliliter). Thereafter, 10 N solution of sodium hydroxide (0.2milliliter) was added thereto and stirred. After 10 N solution of sodiumhydroxide (0.4 milliliter) was further added thereto, the solution wasplaced at room temperature to obtain white needle form crystals. Thecrystals were filtered, washed with water, and then dried in air (0.8 g,yield 80%). Example 5 Preparation of the crystalline Form V of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide

The crystalline Form IV prepared in Example 3 or 4 was dried undervacuum in the presence of P₂O₅ for one day to obtain the titledcrystalline Form V.

Example 6 Preparation of the crystalline Form VI (1) of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide

The crystalline Form V prepared in Example 5 was placed for one day atthe relative humidity of 53% to obtain the titled crystalline Form VI.

Example 7 Preparation of the crystalline Form VI (2) of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide

The crystalline Form V prepared in Example 5 was placed in a glasscontainer, and nitrogen saturated with water was passed through thecontainer for one hour to obtain the titled crystalline Form VI.

Example 8 Preparation of the crystalline Form 1 of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide

Water was added to all the crystalline forms except the crystalline FormI and the mixture was stirred for one hour or more to obtain the titledcrystalline Form I.

Comparative Example 1 Preparation of the amorphous form of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide

The crystalline Form III obtained at Example 2 was dried under vacuum inthe presence of P₂O₅ for two days to obtain the titled amorphous form.

Test Example 1 Powder X-ray diffraction test of the free compound of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide

Each 40 mg of the crystalline Form 1 and the crystalline Form IVprepared in Example 8 and Example 3 or 4 was thinly coated onto a sampleholder, and thereafter the powder x-ray diffraction test was conductedthereto according to the following conditions. By using Rigaku GeigeflexD/max-III C apparatus, the test was conducted at 35 kV, 20 mA.

-   -   Scan speed(2E) 5°/minute    -   Sampling time 0.03 sec    -   Scan mode: continuous    -   Cu-target (Ni filter)

The results of the powder X-ray diffraction test to the crystalline FormI and Form IV are shown in FIGS. 1 and 4. The positions of peaks shownin the above figures are listed at Tables 1 and 2. As shown in eachresult, each crystalline form has different crystallinity. TABLE 1 Peaksof the powder X-ray diffraction of the crystalline Form I of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamidepeak 2θ 7.319 7.81 9.117 10.02 10.808 11.397 13.01 13.732 14.192 15.34616.05 16.539 18.003 19.425 20.01 21.111 21.832 22.226 22.802 23.21224.368 24.781 25.289 26.129 26.698 27.257 27.568 28.802 29.632 30.867

TABLE 2 Peaks of the powder X-ray diffraction of the crystalline Form IVof (2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamidepeak 2θ 8.923 9.966 10.845 11.727 12.395 13.335 13.843 14.778 15.59116.686 17.819 18.364 18.85 19.419 19.871 20.835 21.92 23.06 23.61724.629 25.09 26.017 26.746 27.522 27.872 29.043 30 30.649 31.547

Test Example 2 Powder X-ray diffraction test during moisture absorptionand dehumidification of the crystalline Form I of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide

40 mg of the above crystalline Form I was thinly coated onto a sampleholder. And, immediately after the sample was dried under vacuum in thepresence of P₂O₅, and after the sample was placed for moistureabsorption at each relative humidity of 33%, 53%, 64%, 75%, and 93% fortwo days or more, respectively, the powder X-ray diffraction test wasconducted on the sample according to the conditions represented in aboveTest example 1 to observe change of the crystalline formi duringmoisture absorption. While lowering the relative humidity, the same testwas repeated to observe change of the crystalline form duringdehumidification.

In order to obtain each relative humidity above, as shown in the tablebelow, saturated aqueous solutions of salts were prepared, then placedin a desiccator, and the desiccator was sealed. TABLE 3 RelativeHumidity 33% MgCl₂ saturated aqueous solution Relative Humidity 53%Mg(NO₃)₂.6H₂O saturated aqueous solution Relative Humidity 64% NaNO₂saturated aqueous solution Relative Humidity 75% NaCl saturated aqueoussolution Relative Humidity 93% KNO₃ saturated aqueous solution

The results of the powder X-ray diffraction test of the crystalline FormII exhibited immediately after the vacuum drying to the relativehumidity of 75%, and of the crystalline Form III exhibited at therelative humidity of 64%˜33% during dehumidification are provided inFIGS. 2 and 3, respectively. The positions of peaks shown at the figuresare listed at the following Tables 4 and 5. Each result shows that eachcrystalline form has different crystallinity. TABLE 4 Peaks of thepowder X-ray diffraction of the crystalline Form II of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamidepeak 2θ 7.012 7.822 9.739 10.607 11.43 12.15 13.841 15.17 16.384 17.12217.802 19.198 20.052 20.954 21.882 22.68 23.713 24.837 25.438 25.90226.387 28.046 28.501 28.935 29.304 29.856 30.866 31.405 32.098 33.016

TABLE 5 Peaks of the powder X-ray diffraction of the crystalline FormIII of (2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamidepeak 2θ 7.335 9.09 9.808 10.601 11.203 11.761 13.44 15.245 15.755 19.38920.86 21.629 24.436 26.236 27.159 29.123 29.73 30.763

Test Example 3 Powder X-ray diffraction test during moisture absorptionand dehumidification of the crystalline Form IV of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide

40 mg of the above crystalline Form IV was thinly coated onto a sampleholder. Immediately after the sample was dried under vacuum in thepresence of P₂O₅, and after the sample was placed for moistureabsorption at each relative humidity of 33%, 53%, 64%, 75%, and 93% fortwo days or more, respectively, the powder X-ray diffraction test of thesample was conducted according to the conditions represented in Testexample 1 above to observe change of the crystalline form duringmoisture absorption. While lowering the relative humidity, the same testwas repeated to observe change of the crystalline form duringdehumidification.

In order to obtain each relative humidity above, as shown in Table 3 ofTest example 2, saturated aqueous solutions of salts were prepared andthen placed in a desiccator, and the desiccator was sealed.

The results of the powder X-ray diffraction test of the crystalline FormV exhibited immediately after the vacuum drying and of the crystallineForm VI exhibited after moisture absorption get started are provided inFIGS. 5 and 6, respectively. The positions of peaks shown at the figuresare listed in the following Tables 6 and 7. Each result shows that eachcrystalline form has different crystallinity. TABLE 6 Peaks of thepowder X-ray diffraction of the crystalline Form V of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamidepeak 2θ 8.739 9.878 10.789 11.716 12.451 13.965 14.567 15.368 15.85817.093 17.757 18.296 19.674 20.319 20.799 22.227 23.112 23.742 24.59625.873 26.458 27.502 27.935 28.68 29.358

TABLE 7 Peaks of the powder X-ray diffraction of the crystalline Form VIof (2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamidepeak 2θ 8.042 8.718 10.231 10.78 11.668 12.445 13.56 14.682 15.22215.864 16.5 17.084 17.814 18.698 19.225 19.659 20.327 21.14 22.54123.246 24.656 25.275 25.86 26.636 27.453 28.584 29.147 29.755 30.793

Test example 4 Stress stability test for the amorphous form, and thecrystalline Form I and Form VI

In order to compare physicochemical stability among the crystalline FormVI, the crystalline Form I, and the amorphous form prepared in Examples7, 8, and Comparative Example 1, the stress stability test was conductedby placing their samples at the temperatures of −20° C., 50° C., and 70°C. for 4 weeks. The results are summarized at the following Table 8.TABLE 8 Form I Form VI Amorphous form Color Ivory Ivory Yellow Residualrate −20° C. 99% 101% 96% after 4 weeks  50° C. 99%  99% 96% Residualrate  70° C. 100%  100% 87%

Industrial Applicability

As shown from the above results, the crystalline Form I and Form VIexhibited remarkably superior stability over the amorphous form. Theamorphous form did not show any change in appearance at −20° C. and 50°C., but showed a residual rate of 96% after 4 weeks. At 70° C., theamorphous form showed a residual rate of 87% as well as a change inappearance. Therefore, it can be seen that the crystalline formsaccording to the present invention show superior physicochemicalstability over the amorphous form.

1. Crystalline forms of(2S)-N-5-[amino(imino)methyl]-2-thienylmethyl-1-(2R)-2-[(carboxymethyl)amino]-3,3-diphenylpropanoyl-2-pyrrolidinecarboxamide.nH₂Orepresented by the following Formula (1):

wherein n is the number of combined water per molecule and represents 0,1, 3, 4, 6, or 7.5:
 2. The crystalline forms of claim 1 wherein nrepresents
 1. 3. The crystalline forms of claim 1 or claim 2 whereinX-ray diffraction angles are 13.6°, 14.7°, 23.2°, and 27.5°.
 4. Thecrystalline forms of claim 1 wherein water content is 2 to 9%.
 5. Thecrystalline forms of claim 1 wherein n represents
 4. 6. The crystallineforms of claim 1 or claim 5 wherein X-ray diffraction angles are 7.0°,12.2°, and 19.2°.
 7. The crystalline forms of claim 1 wherein watercontent is 9 to 15%.
 8. The crystalline forms of claim 1 wherein nrepresents 7.5.
 9. The crystalline forms of claim 1 or claim 8 whereinX-ray diffraction angles are 7.3°, 9.1°, 18.0°, and 28.8°.
 10. Thecrystalline forms of claim 1 wherein water content is 16 to 26%.